Cancer is one of the leading causes of death in the developed world, with over one million people diagnosed with cancer and 500,000 deaths per year in the United States alone. Overall it is estimated that more than one in three people will develop some form of cancer during their lifetime. There are more than 200 different types of cancer, four of which—breast, lung, colorectal (CRC) and prostate—account for over half of all new cases (Jemal et al., Cancer J. Clin. 53:5-26 (2003)). CRC is the most common diagnosed cancer in the United States (Jemal et al., CA Cancer J. Clin. 60(5):277-300 (2010)). Chemoprevention using pharmaceutical agents to treat precancerous conditions is widely believed to be a promising strategy to reduce the incidence of and mortality from CRC and other cancer types, especially in individuals at high risk who develop large numbers of precancerous lesions that cannot be effectively removed by colonoscopy or who develop such lesions sporadically that go undetected and become cancerous. Examples of such high risk individuals include those with familial or sporadic adenomatous polyposis, hereditary non-polyposis colon cancer, and inflammatory bowel diseases, including Chrohn's disease and ulcerative colitis. While numerous mutated genes have been shown to be involved in the development of CRC, few molecular targets have been identified that are critically involved in disease initiation and progression and which are suitable drug targets. As such, few drugs have been developed and approved by the FDA to be safe and effective for cancer chemoprevention. The cyclooxygenase-2 selective inhibitor, celecoxib (Celebrex®) is one example that was approved for the treatment of familial adenomatous polyposis, but was recently withdrawn from the market. As such, there is a significant unmet medical need for new drugs to treat or prevent precancerous and cancerous conditions.
Epidemiological studies have shown that non-steroidal anti-inflammatory drugs (NSAIDs) including cyclooxygenase-2 inhibitors (Coxibs) display promising CRC chemopreventive efficacy (Smalley et al., Arch. Intern. Med. 159(2):161-6 (1999) (this is redundant with the 1st and last sentence in this paragraph) and other cancer types. Clinical studies have reported that certain prescription strength NSAIDs such as sulindac (Clinoril®) also have pronounced benefits for individuals with familial adenomatous polyposis (FAP) by causing the regression of precancerous adenomas, reducing both the number and size of such lesions (Giardiello et al., N. Engl. J. Med. 328(18):1313-6 (1993)). COX-2 selective inhibitors (e.g., celecoxib, Celebrex®) have similar benefits, but tend to be less effective and require higher dosages compared with sulindac(Steinbach et al., N. Engl. J. Med. 342(26):1946-52 (2000)). NSAIDs may also be effective for treating advanced stage malignant disease. For example, a clinical trial involving patients with metastatic disease reported that indomethacin (a sulindac analog) extended survival by approximately 9 months (Lundholm et al., Cancer Res. 54(21):5602-6 (1994)). Despite these promising observations, NSAIDs and COX-2 inhibitors are not recommended for cancer chemoprevention because of potentially fatal gastrointestinal, renal and cardiovascular toxicity that result from COX-1 or COX-2 inhibition and suppression of physiologically important prostaglandins (Mukherjee, Biochem. Pharmacol. 63(5):817-21 (2002)).
Still, in view of the strong cancer chemopreventive activity of NSAIDs, increasing efforts have been made to understand the underlying mechanism of action to develop improved drugs that are safer and more efficacious. While the molecular basis for the antineoplastic activity of NSAIDs is commonly attributed to COX-2 inhibition, multiple investigators have concluded that mechanisms other than COX inhibition may be involved (Alberts et al., J. Cell Biochem. Suppl. 22:18-23 (1995); Soh et al., Prog. Exp. Tumor. Res. 37:261-85 (2003); Williams et al., Cancer Res. 60:6045-6051 (2000)). For example, the sulfone metabolite of sulindac has been shown to inhibit tumorigenesis in various rodent models of CRC and other cancer types, despite its inability to inhibit COX (Goluboff et al., Urology 53(2):440-5 (1999); Malkinson et al., Carcinogenesis 19(8):1353-6 (1998); Piazza et al., Cancer Res. 57(14):2909-15 (1997); Thompson et al., Cancer Res. 57(2):267-71 (1997)). Knowledge of the underlying mechanism could lead to the identification of new molecular targets that will provide insight to the discovery of new drugs for cancer intervention and treatment. Studies have shown that the mechanism responsible for the antineoplastic activity of sulindac sulfone (exisulind) involves cyclic guanosine monophosphate phosphodiesterase (cGMP PDE) inhibition (Piazza et al., Cancer Res. 61(10):3961-8 (2001); Thompson et al., Cancer Res. 60(13):3338-42 (2000). More recently, it has been reported that the COX inhibitory sulfide metabolite of sulindac (SS) and other NSAIDs also inhibit cGMP PDE, and this activity is closely associated with their tumor cell growth inhibitory activity (Tinsley et al., Mol. Cancer Ther. 8(12):3331-40 (2009); Tinsley et al., Cancer Prey. Res. (Phila) 3(10):1303-13 (2010); Whitt et al., Cancer Prey. Res. (Phila) 5(6):822-33 (2012); Zhu et al., Curr. Top. Med. Chem. 7(4):437-54 (2007); Tinsley et al., Cancer Prey. Res. (Phila) 4(8):1275-84 (2011)). Other investigators have also suggested a relationship between cGMP elevation and CRC chemoprevention based on several independent lines of evidence (Soh et al., Mol. Carcinog. 47(7):519-25 (2008); Saha et al., J. Appl. Toxicol. 28(4):475-83 (2008); Soh et al., Clin. Cancer Res. 6(10):4136-41 (2000); Kwon et al., Cancer 112(7):1462-70 (2008)).
Phosphodiesterases (PDEs) are a class of intracellular enzymes involved in signal transduction by catalyzing the hydrolysis of the cylic nucleotides, cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphates (cGMP) into their respective, inactive nucleotide monophosphates. The cyclic nucleotides cAMP and cGMP are synthesized by adenylyl and guanylyl cyclases, respectively, and serve as secondary messengers in multiple biochemical pathways that include the activation of cAMP and cGMP-dependent kinases and subsequent phosphorylation of specific proteins that regulate cellular activity and function. For example, cyclic nucleotides in neurons are involved in the acute regulation of synaptic transmission as well as in neuronal differentiation and survival. The complexity of cyclic nucleotide signaling is reflected by the molecular diversity of the enzymes involved in the synthesis and degradation of cAMP and cGMP. There are at least ten families of adenylyl cyclases, two families of guanylyl cyclases, and eleven families of phosphodiesterases. Furthermore, different types of neurons are known to express multiple isozymes of each of these classes, and there is evidence for compartmentalization and specificity of function for different isozymes within a given cell type.
On the basis of substrate specificity, the PDE families can be further classified into three groups: i) the cAMP-PDEs (PDE4, PDE7, PDE8), ii) the cGMP-PDEs (PDE5, PDE6 and PDE9), and iii) the dual-substrate PDEs (PDE1, PDE2, PDE3, PDE10 and PDE11). Furthermore, PDEs are differentially expressed throughout the organism and are generally believed to have distinct physiological functions. As a result of these distinct enzymatic activities and complex tissue localization patterns, different PDE isozyme families can serve as specific targets for distinct therapeutic indications. Furthermore, compounds that can selectively inhibit distinct PDE families or isozymes may offer particular tissue specificity, greater efficacy, and fewer side effects.
Although sulindac can inhibit multiple cGMP degrading isozymes, previous studies have reported that inhibition of the cGMP-specific PDE5 isozyme is closely associated with its anticancer activity (Tinsley et al., Mol. Cancer Ther. 8(12):3331-40 (2009); Tinsley et al., Cancer Prev. Res. (Phila) 3(10):1303-13 (2010); Whitt et al., Cancer Prev. Res. (Phila) 5(6):822-33 (2012); Tinsley et al., Cancer Prev. Res. (Phila) 4(8):1275-84 (2011)). However, highly potent PDE5 selective inhibitors like sildenafil inhibit tumor cell growth with low potency at concentrations that significantly exceed the concentration required for PDE5 inhibition. As such, there is the possibility that additional PDE isozymes may be involved.
PDE10, also known in the art as PDE10A, PDE10A1, or PDE10A2, is identified as a unique PDE isozyme family based on primary amino acid sequence and distinct enzymatic activity. The PDE10 family of polypeptides shows a lower degree of sequence homology as compared to previously identified PDE families and has been reported to be insensitive to certain inhibitors that are known to be specific for other PDE families. PDE10 was first discovered in 1999 (Loughney et al., Gene 234(1):109-17 (1999); Fujishige et al., Eur. J. Biochem. 266(3):1118-27 (1999); Fujishige et al., J. Biol. Chem. 274(26):18438-45 (1999); Soderling et al., Proc. Natl. Acad. Sci. USA 96(12):7071-6 (1999)). Scientific literature has reported that PDE10 is highly expressed in brain striatum, testes, and thyroid but is not or has low expression in most other peripheral tissues. See, Seeger et al., Brain Res. 985(2):113-26 (2003); Kotera et al., J. Biol. Chem. 279(6):4366-75 (2004); Xie et al., Neuroscience 139(2):597-607 (2006); Coskran et al., J. Histochem. Cytochem. 54(11):1205-13 (2006). The high expression of PDE10 in the striatum has suggested a role of this isozyme in various neurological diseases including Parkinson's disease, Huntington's disease, and schizophrenia.